TY - JOUR
T1 - Ultrafast enzymatic digestion of proteins by microdroplet mass spectrometry
AU - Zhong, Xiaoqin
AU - Chen, Hao
AU - Zare, Richard N.
N1 - Funding Information:
This work was supported by the Scientific Research Startup Foundation (IDH1615113) of Fudan University, NSF (CHE-1915878), and key postdoc funding (KLH1615199).
Publisher Copyright:
© 2020, The Author(s).
PY - 2020/12/1
Y1 - 2020/12/1
N2 - Enzymatic digestion for protein sequencing usually requires much time, and does not always result in high sequence coverage. Here we report the use of aqueous microdroplets to accelerate enzymatic reactions and, in particular, to improve protein sequencing. When a room temperature aqueous solution containing 10 µM myoglobin and 5 µg mL−1 trypsin is electrosonically sprayed (−3 kV) from a homemade setup to produce tiny (∼9 µm) microdroplets, we obtain 100% sequence coverage in less than 1 ms of digestion time, in sharp contrast to 60% coverage achieved by incubating the same solution at 37 °C for 14 h followed by analysis with a commercial electrospray ionization source that produces larger (∼60 µm) droplets. We also confirm the sequence of the therapeutic antibody trastuzumab (∼148 kDa), with a sequence coverage of 100% for light chains and 85% for heavy chains, demonstrating the practical utility of microdroplets in drug development.
AB - Enzymatic digestion for protein sequencing usually requires much time, and does not always result in high sequence coverage. Here we report the use of aqueous microdroplets to accelerate enzymatic reactions and, in particular, to improve protein sequencing. When a room temperature aqueous solution containing 10 µM myoglobin and 5 µg mL−1 trypsin is electrosonically sprayed (−3 kV) from a homemade setup to produce tiny (∼9 µm) microdroplets, we obtain 100% sequence coverage in less than 1 ms of digestion time, in sharp contrast to 60% coverage achieved by incubating the same solution at 37 °C for 14 h followed by analysis with a commercial electrospray ionization source that produces larger (∼60 µm) droplets. We also confirm the sequence of the therapeutic antibody trastuzumab (∼148 kDa), with a sequence coverage of 100% for light chains and 85% for heavy chains, demonstrating the practical utility of microdroplets in drug development.
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U2 - 10.1038/s41467-020-14877-x
DO - 10.1038/s41467-020-14877-x
M3 - Article
C2 - 32103000
AN - SCOPUS:85080102070
SN - 2041-1723
VL - 11
JO - Nature communications
JF - Nature communications
IS - 1
M1 - 1049
ER -